Summary: This study represents an important step in our efforts to reconstitute a ‘minimal mitotic spindle’ with purified proteins. The biophysical and structural characterization of the key non-motor microtubule-associated proteins (MAPs) required for cell division lags far behind that of motor proteins (e.g. kinesins). To fill this knowledge gap and to characterize the proteins needed for our reconstitution assays, we focused on PRC1. This non-motor MAP, or its homologs, is needed for normal cell growth in plants and fungi. In metazoans, PRC1 is needed for successful cell division. We combined X-ray crystallography, electron microscopy, and TIRF (total internal reflection fluorescence) microscopy approaches to analyze PRC1’s functions. We showed that PRC1’s microtubule binding involves two domains: a structured domain with a spectrin-fold, a motif not previously known to mediate interactions with microtubules, and an unstructured positively charged domain. We find that single PRC1 dimers are flexible when bound to one microtubule, a feature that may help the protein find a second microtubule in the cytoplasm to increase crosslinking efficiency. In contrast, PRC1 adopts a rigid conformation between two filaments, as would be needed to achieve specificity in filament crosslinking geometry. TIRF microscopy-based assays revealed that PRC1 autonomously and dynamically tracks microtubule overlap regions, generating crosslinks that are compliant and don’t block relative filament movements. These data suggest that in self-organizing filament networks, PRC1 could ‘mark’ regions where microtubules have a specific geometry. It would allow the network to remain dynamic, but recruit kinases and other motor proteins to modulate filament position and stability. Together, our findings suggest how microtubule-binding proteins, such as PRC1, recognize nanometer-scale features and thereby transmit positional information to kinases, such as Aurora, that can then generate micron-scale spatial phosphorylation gradients to control the assembly and function of dynamic architectures in the cell.